Internal death programs play significant roles in many diseases. Pathogenic effects can result from inefficient cell death or from inappropriate or excessive death such as that caused by the human immunodeficiency virus (HIV) during AIDS or the SAR-CoV virus during SARS. In this project, we are taking a multifaceted approach to studying molecular mechanisms of both apoptotic and nonapoptotic death programs in lymphocytes as well as other cell types. A major focus of our investigations are death-inducing cell surface receptors in the tumor necrosis factor receptor (TNFR) superfamily such as TNFR1 and CD95/Fas/APO-1. Both receptors play an important role in stimulating both apoptotic and nonapoptotic death of cells principally in immune processes. Little is known about how these alternative death pathways are entrained to receptor signaling. Interestingly, both receptors can have effects beside death such as the induction of transcription factors. We are trying to understand how these receptors stimulate the intracellular machinery that causes cell death in preference to other cellular outcomes. We have discovered that inhibition of caspase-8 in non-lymphoid cells can lead to another form of cell death exhibiting particular cytoplasmic double membrane structures called autophagy. Although initially controversial, several labs have now shown that this form of death is particularly important for the demise of tumor cells by chemotherapeutic agents. We have now shown that the mechanism of autophagic death program is selective degradation of catalase which leads to a marked overaccumulation of reactive oxygen species leading to cellular damage and death. Furthermore, we have focused on genes that play key roles in this process of death. Autophagy is an evolutionarily conserved process from humans to yeast by which cytoplasmic proteins and organelles are catabolized but very little was known about results at the end of autophagy when cells were selecting between autophagic cell death and survival. During starvation, the protein TOR (target of rapamycin), a nutrient-responsive kinase that controls cellular metabolism, is shut off, and autophagy is activated. Double-membrane autophagosomes sequester intracellular components and then fuse with lysosomes to form autolysosomes, which to catabolize their contents to regenerate nutrients. Ourpresent understanding of autophagy is that it terminates with cargo degradation within autolysosomes, but how autophagy is controlled by nutrients and the subsequent fate of the autolysosome were unknown. We discovered that mTOR signalling in mammalian cells is inhibited during initiation of autophagy, but reactivated during extended starvation. Reactivation of mTOR depends on the degradation of autolysosomal products and release of nutrients. mTOR activity in turn terminates autophagy and stimulates impressive proto-lysosomal tubules and vesicles that extrude from autolysosomes and ultimately mature into functional lysosomes. This process, that we term autophagic lysosome reforation (ALR), restores the full complement of lysosomes in the cell. This evolutionarily conserved cycle in autophagy governs nutrient sensing and lysosome homeostasis during starvation. In parallel, we are exploring how the regulation of cellular death programs may play a role in cytopathicity associated with virus infections in AIDS and SARS. In particular, a critical effect in the onset of AIDS following infection with HIV is the death of T lymphocytes caused by the virus. We previously found that this death process was necrotic rather than apoptotic. In 2012, we carried out a study of the two major cytopathic factors in human immunodeficiency virus type 1 (HIV-1), the accessory proteins viral infectivity factor (Vif) and viral protein R (Vpr), that inhibit cell-cycle progression at the G2 phase of the cell cycle which led to cause necrotic cell death. Although Vpr-induced blockade and the associated T-cell death have been well studied, the molecular mechanism of G2 arrest by Vif remains undefined. To elucidate how Vif kills the cell by inducing G2 arrest, we infected synchronized Jurkat human T-cells and examined the effect of Vif on the activation of Cdk1 and CyclinB1, the chief cell-cycle factors for the G2 to mitosis phase transition. We found that the characteristic dephosphorylation of an inhibitory phosphate on Cdk1 did not occur in infected cells expressing Vif. In addition, the nuclear translocation of Cdk1 and CyclinB1 was impaired. Finally, Vif-induced cell cycle arrest and cytotoxicity was correlated with proviral expression of Vif. we concluded that Vif impairs mitotic progression and causes fatal cell cycle disruption by interfering with Cdk1-CyclinB1 activation. In 2012, we also found that during autophagy, multiple lysosomes fuse with an autophagosome to form an autolysosome in which cytoplasmic components are sequestered and degraded by lysosomal hydrolases which releases the products into the cytosol via lysosomal efflux permeases. Following starvation-induced autophagy, lysosome homeostasis is restored by autophagic lysosome reformation (ALR) requiring activation of the target of rapamycin (TOR) kinase. Spinster (Spin) encodes a putative lysosomal efflux permease similar to a sugar transporter. Spin mutants accumulate lysosomal carbohydrates and generate enlarged lysosomes in Drosophila and in human cells. We also demonstrated that spin is crucial for mTOR reactivation and lysosome reformation during prolonged starvation. Finally, we demonstrate that the sugar transporter activity of Spin is essential for ALR. These results provide a detailed molecular insight into how lysosome biogenesis proceeds during starvation and potentially during other autophagy-inducing conditions. We also conjecture that this mechanism may underlie normal lysosome biogenesis even in nutrient replete conditions.